This project is broadly concerned with an elucidation of the structural, physicochemical, and reactivity properties of the three currently recognized types of active sites, (Fe(S-Cys)4), (Fe2S2(s-Cys)4), and (Fe4S4(S-Cys)4), found in iron sulfur proteins and enzymes. The basic approach to this problem is by synthesis of lower molecular weight analogues of these sites followed by investigation of the preceding properties and detailed comparisons of related properties (where known) of proteins an enzymes. Substantial progress has already been made under NIH support, resulting in synthesis and detailed characterization of the three structural types of sites in five of the seven physiologically significant oxidation levels. The following research is proposed in the coming grant year: (1) synthesis of (Fe2S2(SR)4) 3 minus and (Fe4S4(SR)4) 1 minus (the two remaining analogues of physiological oxidation levels) using a tetracysteinyl dodecapeptide as a stabilizing ligand; (2) continued development of the 19F nmr core extrusion technique for the identification of Fe2Se and Fe4S4 sites in proteins, and application of the FeMo proteins of nitrogenase and membrane-bound proteins in subchloroplast photosynthetic particles; (3) elucidation of reactions of the hydrogenase and nitrogenase inhibitors CO and No with (Fe4S4(SR)4) 3 minus, 2 minus , analogues of sites present in these enzymes; (4) synthesis and structural characterization of (Fe4S4(SR)4) 4 minus, a species not likely to be subject to structural distortion; (5) development of (Fe4S4(SR)4) 2n minus n as models for multi-electron transfer proteins.